COVID-19 severity and in-hospital mortality in an area with high HIV prevalence

Background HIV infection causes immune dysregulation affecting T-cell and monocyte function, which may alter coronavirus disease 2019 (COVID-19) pathophysiology. Objectives We investigated the associations among clinical phenotypes, laboratory biomarkers, and hospitalisation outcomes in a cohort of people hospitalised with COVID-19 in a high HIV prevalence area. Method We conducted a prospective observational cohort study in Tshwane, South Africa. Respiratory disease severity was quantified using the respiratory oxygenation score. Analysed biomarkers included inflammatory and coagulation biomarkers, CD4 T-cell counts, and HIV-1 viral loads (HIVVL). Results The analysis included 558 patients, of whom 21.7% died during admission. The mean age was 54 years. A total of 82 participants were HIV-positive. People living with HIV (PLWH) were younger (mean age 46 years) than HIV-negative people; most were on antiretroviral treatment with a suppressed HIVVL (72%) and the median CD4 count was 159 (interquartile range: 66–397) cells/µL. After adjusting for age, HIV was not associated with increased risk of mortality during hospitalisation (age-adjusted hazard ratio = 1.1, 95% confidence interval: 0.6–2.0). Inflammatory biomarker levels were similar in PLWH and HIV-negative patients. Detectable HIVVL was associated with less severe respiratory disease. In PLWH, mortality was associated with higher levels of inflammatory biomarkers. Opportunistic infections, and other risk factors for severe COVID-19, were common in PLWH who died. Conclusion PLWH were not at increased risk of mortality and those with detectable HIVVL had less severe respiratory disease than those with suppressed HIVVL. What this study adds This study advances our understanding of severe COVID-19 in PLWH.


Introduction
People living with HIV (PLWH) have an increased risk of mortality from infection with respiratory viruses, including influenza and human metapneumovirus. 1,2 Many studies have reported that PLWH, especially those not on antiretroviral treatment (ART) and with a detectable HIV-1 viral load (HIVVL), have a higher risk of coronavirus disease 2019 (COVID-19)-related in-hospital mortality. 3,4,5 However, observational cohort studies of hospitalised patients with COVID-19 have reported that PLWH had lower oxygen requirements during their admission. In these studies, patients with detectable HIVVL had lower relative risk of intubation than PLWH with suppressed HIVVL. 6,7 In an observational study, PLWH had similar outcomes after initiation of high-flow nasal oxygen or ventilatory support. 8 There is still some uncertainty as to whether COVID-19 immunopathology and clinical phenotypes are altered by HIV coinfection.
Current evidence suggests that severe COVID-19 is associated with dysregulation of the monocyte-macrophage response, defective T-cell responses, elevated inflammatory cytokines, and hyperactivated neutrophils which culminate in ongoing, inappropriate systemic inflammation which damages pulmonary and other tissues. 9,10,11,12 People hospitalised with COVID-19, particularly with the more severe spectrum of disease, can develop acute respiratory distress http://www.sajhivmed.org.za Open Access syndrome (ARDS), which is associated with systemic inflammation. 13 Established markers of respiratory disease severity in COVID-19 include respiratory oxygenation (ROX) scores and partial pressure of oxygen (PaO 2 )/fraction of inspired oxygen (FIO 2 ) ratios. 8,13,14 Antiviral therapy in early COVID-19 leads to lower risk of hospitalisation and mortality. 15,16,17 In addition, anti-inflammatories with either broad and non-specific targets, like high-dose corticosteroids, or targeted, like the interleukin 6 inhibitor tocilizumab, also lead to reduced risk of in-hospital mortality. 18 HIV infection has strong effects on cellular immune phenotypes and function, affecting T and B lymphocytes, and monocytes -all of which are implicated in COVID-19 pathophysiology. 19,20,21 Recently, more evidence has emerged to assess the effect of HIV on cellular immune responses on COVID-19. HIV coinfection does not appear to alter severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) CD4+ function or phenotypes, but is associated with reduced CXCR3 expression on CD8+ T-cells. 22,23 Higher HIVVLs are also associated with increased expression of activation markers on CD8+ T-cells in COVID-19, which may alter disease phenotypes. People living with HIV mount similar SARS-CoV-2-specific antibody responses in acute COVID-19 to HIV-negative people. 24 HIV viraemia alters monocyte subpopulation phenotypes, reducing CCR2 and CX3CR1 expression, which may affect their ability to move from blood into tissue. In COVID-19 this may reduce pulmonary inflammation. 21 The majority of PLWH analysed in larger cohorts or systematic reviews were on ART, with suppressed HIVVL and high CD4 T-cell counts. 3,4,25 Consequently, the effect of HIV viraemia and associated immunological changes on the clinical manifestations of COVID-19 remain poorly described. Uncertainty also exists as to the extent to which comorbidities in PLWH, such as hypertension, diabetes, or opportunistic infections, contribute to the increased risk of morbidity and mortality.
We investigated whether HIV infection is associated with COVID-19 severity, differences in routinely collected laboratory biomarkers, and in-hospital mortality in a wellcharacterised clinical cohort of patients admitted with COVID-19, in a setting with a high HIV prevalence.

Cohort description
We conducted a prospective, single centre, observational cohort study of patients admitted to the Tshwane District Hospital complex (TDH) from April 2020 to November 2020. Admission vital sign data were taken as the worst score within a 48 h window around the date of admission (admission ± 24 h). The ROX score was calculated for participants with admission vital data (Online Appendix 1 Figure 1-A1). 14 The ROX score is a continuous variable which estimates respiratory disease severity by creating a composite score considering the supplemental oxygen concentration, peripheral oxygen saturation, and respiratory rate.
Laboratory biomarker data were extracted from the South African National Health Laboratory Services (NHLS) online data warehouse. We analysed haematology panels (full blood count; differential white cell counts including absolute neutrophil count [

Cohort description
A total of 558 patient records were analysed ( There was no difference in the proportion of symptoms reported at admission, with cough and dyspnoea the most common ( Figure 2) (chi-squared P > 0.05 for all comparisons).

Respiratory oxygenation scores and laboratory biomarkers are associated with COVID-19 severity and mortality
The median ROX score at admission was 8.2 (IQR: 4.8-16.7). Increasing age was associated with lower ROX scores at  admission (correlation of age and ROX score: rho = -0.2, P < 0.001), and higher DDIM and inflammatory biomarker levels ( Figure 2a and Figure 2b). ROX scores, inflammatory biomarkers and DDIM levels showed significant collinearity with each other (Figure 2b).
A total of 85 patients required HC as their highest level of supportive care and 48 patients were admitted to ICU. Length of hospitalisation was longer for those admitted to ICU than the general wards (median 16 days vs 5 days, P < 0.001). In addition, patients admitted to ICU had more severe disease -ROX scores were lower, ANC, PCT, DDIM and ALT levels were significantly higher when compared to patients admitted to HC or general wards (Online Appendix 1 Table 1-A1). Consequently, higher ROX scores, ANC, NLR and PCT levels predicted admission to ICU with AUROC > 0.7. ROX scores below 4.5 had the highest specificity at 82% (sensitivity 62%), and PCT above 0.13 the highest sensitivity at 94% (specificity 57%). The mortality rate was 57% for patients admitted to ICU, 24% for HC, and 17% for general ward admissions (FET P < 0.001 for ICU vs HC or General wards).
A total of 121 (21.7%) patients died during admission.
Increasing age was associated with increased mortality during admission (Figure 2c). A ROX score below six at admission was associated with a twofold increase in mortality compared to higher ROX scores (age-adjusted hazard ratio [aHR] = 2.1, 95% CI: 1.2-3.6, P = 0.01). The tertiles grouping the highest levels of creatinine, CRP, ferritin, NLR and DDIM were also associated with increased mortality (Figure 3c).

COVID-19 severity in people living with HIV
Respiratory rates were similar between PLWH and HIVnegative patients; however, PLWH needed less oxygen at admission, and had higher peripheral oxygen saturation readings. Respiratory oxygen scores were higher in PLWH, but the difference was not statistically significant (Table 2). There was also no significant difference in NLR, CRP, ferritin, PCT, DDIM, ALT, or haemoglobin A1c (HbA1c) between PLWH and HIV-negative patients. Creatinine levels were slightly lower in PLWH, when compared to HIV-negative patients. These associations were unchanged in linear regression models which adjusted for age differences in HIV-negative patients and PLWH. People living with HIV were as likely as HIV-negative patients to be admitted to ICU -3.7% of PLWH versus 9.5% HIVnegative patients (FET P = 0.2).

Markers of disease severity in people living with HIV, stratified by CD4 count and HIV-1 viral load
In PLWH, a CD4 count below 200 cells/µL was associated with lower odds of having hypertension, diabetes or being on ART (Online Appendix 1 Table 2-A1). People living with HIV with CD4 counts below 200 cells/µL had higher HIVVLs, NLRs and DDIM levels ( Figure 3a) and were more likely to have TB (25% vs 4%, P = 0.06) when compared to PLWH with higher CD4 counts. Respiratory oxygen scores were equivalent between those with higher and lower CD4 counts (Figure 3a). Admission rates to HC or ICU were equivalent between PLWH with CD4 counts above or below 200 cells/µL.   Figure 3c). None of the PLWH with detectable HIVVLs was admitted to HC or ICU.

Hazard raƟos for in-hospital mortality
Age-adjusted hazard raƟos (P = 0.06), CVD (P < 0.001), or CKD (P < 0.001). Respiratory disease severity was significantly worse at admission in HIVnegative patients who died versus those who survived (ROX score: 4.8 vs 9.5, MW P < 0.001). Mortality in the HIV-negative patients was also associated with higher levels of laboratory biomarkers, higher ANC, lower ALC and higher creatinine levels (MW P < 0.001).
A total of 12/82 (14.8%) PLWH died during admission. People living with HIV who died all had significant HIVrelated comorbidities, or other risk factors for COVID-19related mortality (Table 4). Their hospital stay was shorter than those who survived (median of 3 days vs 7 days), and HIVVL and CD4 counts were not associated with inhospital mortality in univariate analyses (logrank P > 0.05) ( Figure 3b & Figure 3d). Creatinine, CRP, PCT and DDIM levels were higher at admission in PLWH who died compared to those who survived (P < 0.05 for all comparisons). When compared to HIV-negative patients who died, PLWH who died had higher DDIM levels at admission (

Discussion
We report a detailed analysis of clinical phenotypes of COVID-19 in hospitalised patients, with and without HIV, and their association with laboratory biomarkers. ROX, respiratory oxygenation; NLR, neutrophil-to-lymphocyte ratio; CRP, C-reactive protein; DDIM, D-dimer; HIV VL, human immunodeficiency virus viral load; ns, not significant. *, P < 0.05; **, P < 0.01 Days hospitalisaƟon 15 3 Therefore, it is possible that the association between respiratory disease severity and HIVVL was confounded by baseline differences in age and comorbidities in PLWH, and not necessarily because of HIV viraemia.
Effective ART suppresses viral replication and reverses much of the immunopathology of HIV, but T-cell and monocyte phenotypes, as well as levels of systemic immune activation, remain altered for years afterwards. 21,35 Monocytes and cytokines involved in monocyte trafficking are central to the pathophysiology of COVID-19. 11 Monocytes are recruited to tissue by the interaction of their CCR2 receptor and its ligand CCL2, which is dysregulated in severe COVID-19. 9 HIV infection decreases CCR2 expression on monocytes, and this is reversed with suppressive ART. 21   The sample size of PLWH who died was small, and it is therefore difficult to draw statistically supported conclusions on these observations. Many of the larger studies which reported that HIV infection is associated with increased risk of severe COVID-19 and related mortality included a higher proportion of men in their analysis than our study. 3,36,37,38 It is possible that sex and HIV interact with COVID-19 to alter disease phenotypes. Neutrophils play an important role in COVID-19 related immunothrombosis, and neutrophils isolated from women have greater inflammatory responses to interferon, which may allow for better innate immune antiviral response. 39 Men with severe COVID-19 have altered kynurenic acid metabolism which is associated weaker T-cell responses. 40 Further research should be undertaken to investigate the interaction of sex, age, and HIV infection on immune responses.
This analysis has several limitations: We analysed records for patients admitted to a single tertiary academic medical centre in an urban area; many patients are sent there by referral which may bias admission towards those with more severe disease. This cohort's prevalence of HIV infection was within the range for other reported estimates and is likely to be broadly representative of similar hospital cohorts in South Africa. 6,41,42 Approximately one-third of PLWH in this study did not have HIVVLs measured which may have biased the comparison between detectable versus undetectable HIVVL participants. Strengths of our study include prospective data collection, a validated method for quantifying respiratory disease severity on a continuum, many patients with laboratory biomarkers during admission, and PLWH well characterised in terms of other comorbidities. We have generated several hypotheses related to HIV and COVID-19 for future exploration.
This study shows that PLWH who were hospitalised with COVID-19 did not have significantly different in-hospital mortality rates, levels of inflammatory biomarkers or respiratory disease severity. People living with HIV who died often had other risk factors for COVID-19-associated mortality, or AIDS-defining illness.

Funding information
This research received no specific grant from any funding agency in the public, commercial or not-for-profit sectors.

Data availability
Complete individual patient data and the analysis code in R are available to researchers on reasonable request from the corresponding author, M.T.B. †, ROX score at admission; ‡, TB treatment started on empirical grounds; §, Anti-nuclear antibody positive, with bicytopenia, rash and joint pain -admitted for haemoptysis and developed nosocomial SARS-CoV-2 infection.